Heteroalicyclic carboxamidines as inhibitors of inducible nitric oxide synthase; the identification of (2R)-2-pyrrolidinecarboxamidine as a potent and selective haem-co-ordinating inhibitor.

Heteroalicyclic carboxamidines were synthesised and evaluated as inhibitors of nitric oxide synthases. (2R)-2-Pyrrolidinecarboxamidine, in particular, was shown to be a highly potent in vitro (IC(50)=0.12 µM) and selective iNOS inhibitor (>100-fold vs both eNOS and nNOS), with probable binding to the key anchoring glutamate residue and co-ordination to the haem iron.

GW274150 and GW273629 are potent and highly selective inhibitors of inducible nitric oxide synthase in vitro and in vivo.

1 GW274150 ([2-[(1-iminoethyl) amino]ethyl]-L-homocysteine) and GW273629 (3-[[2-[(1-iminoethyl)amino]ethyl]sulphonyl]-L-alanine) are potent, time-dependent, highly selective inhibitors of human inducible nitric oxide synthase (iNOS) vs endothelial NOS (eNOS) (>100-fold) or neuronal NOS (nNOS) (>80-fold). GW274150 and GW273629 are arginine competitive, NADPH-dependent inhibitors of human iNOS with steady state K(d) values of <40 and <90 nM, respectively. 2 GW274150 and GW273629 inhibited intracellular iNOS in J774 cells in a time-dependent manner, reaching IC(50) values of 0.2+/-0.04 and 1.3+/-0.16 microM, respectively. They were also acutely selective in intact rat tissues: GW274150 was >260-fold and 219-fold selective for iNOS against eNOS and nNOS, respectively, while GW273629 was >150-fold and 365-fold selective for iNOS against eNOS and nNOS, respectively. 3 The pharmacokinetic profile of GW274150 was biphasic in healthy rats and mice with a terminal half-life of approximately 6 h. That of GW273629 was also biphasic in rats, producing a terminal half-life of approximately 3 h. In mice however, elimination of GW273629 appeared monophasic and more rapid (approximately 10 min). Both compounds show a high oral bioavailability (>90%) in rats and mice. 4 GW274150 was effective in inhibiting LPS-induced plasma NO(x) levels in mice with an ED(50) of 3.2+/-0.7 mg kg(-1) after 14 h intraperitoneally (i.p.) and 3.8+/-1.5 mg kg(-1) after 14 h when administered orally. GW273629 showed shorter-lived effects on plasma NO(x) and an ED(50) of 9+/-2 mg kg(-1) after 2 h when administered i.p. 5 The effects of GW274150 and GW273629 in vivo were consistent with high selectivity for iNOS, as these inhibitors were of low potency against nNOS in the rat cerebellum and did not cause significant effects on blood pressure in instrumented mice.

Antagonism of human CC-chemokine receptor 4 can be achieved through three distinct binding sites on the receptor.

Chemokine receptor antagonists appear to access two distinct binding sites on different members of this receptor family. One class of CCR4 antagonists has been suggested to bind to a site accessible from the cytoplasm while a second class did not bind to this site. In this report, we demonstrate that antagonists representing a variety of structural classes bind to two distinct allosteric sites on CCR4. The effects of pairs of low-molecular weight and/or chemokine CCR4 antagonists were evaluated on CCL17- and CCL22-induced responses of human CCR4(+) T cells. This provided an initial grouping of the antagonists into sets which appeared to bind to distinct binding sites. Binding studies were then performed with radioligands from each set to confirm these groupings. Some novel receptor theory was developed to allow the interpretation of the effects of the antagonist combinations. The theory indicates that, generally, the concentration-ratio of a pair of competing allosteric modulators is maximally the sum of their individual effects while that of two modulators acting at different sites is likely to be greater than their sum. The low-molecular weight antagonists could be grouped into two sets on the basis of the functional and binding experiments. The antagonistic chemokines formed a third set whose behaviour was consistent with that of simple competitive antagonists. These studies indicate that there are two allosteric regulatory sites on CCR4.

Characterization of selective Calcium-Release Activated Calcium channel blockers in mast cells and T-cells from human, rat, mouse and guinea-pig preparations.

Loss of function mutations in the two key proteins which constitute Calcium-Release Activated Calcium (CRAC) channels demonstrate the critical role of this ion channel in immune cell function. The aim of this study was to demonstrate that inhibition of immune cell activation could be achieved with highly selective inhibitors of CRAC channels in vitro using cell preparations from human, rat, mouse and guinea-pig. Two selective small molecule blockers of CRAC channels; GSK-5498A and GSK-7975A were tested to demonstrate their ability to inhibit mediator release from mast cells, and pro-inflammatory cytokine release from T-cells in a variety of species. Both GSK-5498A and GSK-7975A completely inhibited calcium influx through CRAC channels. This led to inhibition of the release of mast cell mediators and T-cell cytokines from multiple human and rat preparations. Mast cells from guinea-pig and mouse preparations were not inhibited by GSK-5498A or GSK-7975A; however cytokine release was fully blocked from T-cells in a mouse preparation. GSK-5498A and GSK-7975A confirm the critical role of CRAC channels in human mast cell and T-cell function, and that inhibition can be achieved in vitro. The rat displays a similar pharmacology to human, promoting this species for future in vivo research with this series of molecules. Together these observations provide a critical forward step in the identification of CRAC blockers suitable for clinical development in the treatment of inflammatory disorders.

The discovery of phthalazinone-based human H1 and H3 single-ligand antagonists suitable for intranasal administration for the treatment of allergic rhinitis.

A series of potent phthalazinone-based human H(1) and H(3) bivalent histamine receptor antagonists, suitable for intranasal administration for the potential treatment of allergic rhinitis, were identified. Blockade of H(3) receptors is thought to improve efficacy on nasal congestion, a symptom of allergic rhinitis that is currently not treated by current antihistamines. Two analogues (56a and 56b) had slightly lower H(1) potency (pA(2) 9.1 and 8.9, respectively, vs 9.7 for the clinical gold-standard azelastine, and H(3) potency (pK(i) 9.6 and 9.5, respectively, vs 6.8 for azelastine). Compound 56a had longer duration of action than azelastine, low brain penetration, and low oral bioavailability, which coupled with the predicted low clinical dose, should limit the potential of engaging CNS-related side-effects associated with H(1) or H(3) antagonism.

Randomized clinical trial comparing 2 support surfaces: results of the Prevention of Pressure Ulcers Study.

OBJECTIVE: To determine whether a viscoelastic polymer (energy absorbing) foam mattress was superior to a standard hospital mattress for pressure ulcer prevention and to analyze the cost-effectiveness in comparison with standard hospital mattresses. DESIGN: Unblinded, randomized, prospective trial. SETTING: Elderly acute care, rehabilitation, and orthopedic wards at 3 hospitals in the United Kingdom. PARTICIPANTS: 1168 patients at risk of developing pressure ulcers (Waterlow score, 15 to 20), with a median age of 83 years (25th to 75th percentile range, 79-87). INTERVENTIONS: Participants were allocated to either the experimental equipment (CONFOR-Med mattress/cushion combination) or a standard mattress/cushion combination; all were given standard nursing care. Pressure areas were observed daily. MAIN OUTCOME MEASURE: Development of nonblanching erythema. RESULTS: A significant decrease in the incidence of blanching erythema (26.3% to 19.9%; P =.004) and a nonsignificant decrease in the incidence of nonblanching erythema occurred in participants allocated to the experimental equipment. However, when the survival curve plots were analyzed at 7 days, both categories showed statistically significant decreases (P =.0015 and P =.042, respectively). Participants on standard equipment had a relative odds ratio of 1.36 (95% confidence interval [CI], 1.10-1.69) for developing blanching erythema or worse and 1.46 (95% CI, 0.90-1.82) for developing nonblanching erythema or worse. To prevent nonblanching erythema, the number needed to treat (NNT) was 41.9 (95% CI, -82.6-15.3). To prevent any erythema (blanching or nonblanching), the NNT was 11.5 (95% CI, 41.6-9.3). Participants with blanching or nonblanching erythema were significantly less mobile than participants with normal skin and more likely to have worsening mobility (P <.001). For participants with similar pressure ulcer status, mattress type was not associated with difference in mobility. CONCLUSIONS: Regardless of prevention routine, pressure ulcers occur. In this study, the experimental equipment showed statistical significance to standard equipment for prevention of blanching erythema; significance was not achieved for nonblanching erythema. Trend and survival analysis show that a larger study is required to determine whether this nonsignificant difference is genuine.